High telomerase activity and high HTRT mRNA expression differentiate pure myxoid and myxoid/round-cell liposarcomas

Molecular markers characterizing the transition of a myxoid to a more round-cell liposarcoma have not been described. To examine whether telomerase activity, hTRT and hTR mRNA expression were associated with tumor progression in myxoid liposarcoma, we investigated a total of 28 myxoid liposarcomas (13 pure myxoid tumors, 14 mixed-type tumors, and 1 pure round-cell variant) from 19 patients. Telomerase activity was detected by using the fluorescent PCR-based TRAP-assay. Expression of hTRT and hTR mRNAs was determined by the semi-quantitative RT-PCR. On the basis of only one tumor sample per patient, telomerase activity was found in 9 of 9 myxoid/round-cell liposarcomas and in 3 of 10 pure myxoid tumors. Elevated hTRT expression was found in 13 of 17 liposarcomas. All telomerase-positive tumors showed hTRT expression, whereas there were 3 cases showing hTRT expression without telomerase activity. HTR mRNA expression was elevated in all 19 liposarcomas. Thus, only the levels of telomerase activity and of hTRT mRNA expression differentiated pure myxoid liposarcoma and myxoid/round-cell liposarcoma (p < 0.003 and p = 0.029, respectively). We believe that high levels of telomerase activity and of hTRT expression are associated with tumor progression from low-grade pure myxoid to higher-grade malignant round-cell liposarcoma, and may consequently represent a useful prognostic marker for this histological sub-type of soft-tissue tumors.     

Telomere lengths and telomerase activity in dog tissues: a potential model system to study human telomere and telomerase biology

Studies on telomere and telomerase biology are fundamental to the understanding of aging and age-related diseases such as cancer. However, human studies have been hindered by differences in telomere biology between humans and the classical murine animal model system. In this paper, we describe basic studies of telomere length and telomerase activity in canine normal and neoplastic tissues and propose the dog as an alternative model system. Briefly, telomere lengths were measured in normal canine peripheral blood mononuclear cells (PBMCs), a range of normal canine tissues, and in a panel of naturally occurring soft tissue tumours by terminal restriction fragment (TRF) analysis. Further, telomerase activity was measured in canine cell lines and multiple canine tissues using a combined polymerase chain reaction/enzyme-linked immunosorbent assay method. TRF analysis in canine PBMCs and tissues demonstrated mean TRF lengths to range between 12 and 23 kbp with heterogeneity in telomere lengths being observed in a range of normal somatic tissues. In soft tissue sarcomas, two subgroups were identified with mean TRFs of 22.2 and 18.2 kbp. Telomerase activity in canine tissue was present in tumour tissue and testis with little or no activity in normal somatic tissues. These results suggest that the dog telomere biology is similar to that in humans and may represent an alternative model system for studying telomere biology and telomerase-targeted anticancer therapies.     

Telomeric recombination in mismatch repair deficient human colon cancer cells after telomerase inhibition

The majority of human malignancies use telomerase to maintain telomere homeostasis. Antitelomerase therapy is therefore a promising approach for a cancer-specific therapy. The alternative lengthening of telomeres pathway (ALT) is a recombination-based, telomerase-independent mechanism of telomere length control. It is widely believed that ALT could be engaged when cancer cells escape from telomerase inhibition. However, no reports exist that would support this concept of therapy resistance. We inhibited telomerase in a human cancer cell line with a mismatch repair defect and observed a telomerase-independent, ALT-like telomere elongation. This is the first report of inducing a telomerase-independent telomere elongation in human cancer cells when telomerase is inhibited, thus describing a novel mechanism of resistance to antitelomerase therapy.     

Changes of telomerase and telomere lengths in paired normal and cancer tissues of breast.

To attain the immortal phenotype, cancer cells must overcome the mitotic clock. Telomerase activity has been identified to be activated in malignant tumors including breast cancer. Telomerase activity was evaluated in 71 breast cancer tissues and paired normal tissues with the TRAP (telomerase repeat amplification protocol) assay. Telomerase activity was calculated and translated into arbitrary units by computer-assisted densitometry with the control of telomerase activity in the 293 control cell line. In 59 paired breast tissues with telomerase activity, terminal restriction fragment (TRF) lengths were measured using Southern blotting. Relative inhibition (RI), the ratio of inhibited telomerase activity in each tumor tissue compared to that of the 293 control cell line after pre-treatment with 150 microg/ml of RNAse A, was measured. Sixty-three of 71 cancer tissues showed telomerase activity (88.7%) with 75.3+/-17.9 units in densitometry, while no telomerase activity was detected in their paired normal tissues. Telomerase activity was correlated to node metastasis (p=0.02) and stage (p=0.005), but not to tumor size or the hormonal receptor status. TRF lengths were 11. 0+/-4.7 kb in 59 tumor tissues and 11.7+/-2.2 kb in paired normal tissues. TRF lengths did not correlate to any of the clinical parameters. However changes of TRF lengths in tumor tissues compared to those of normal tissues correlated to telomerase activity. RI in the tumor tissues was proportional to telomerase activity without RNAse A pre-treatment. In breast cancer, telomerase activity was specific to tumor tissues and increased with tumor progression. Telomerase activity and changes in TRF lengths can be used as guidelines in detecting candidates for the telomerase inhibitor.     

检测人端粒酶活性的端粒酶TRAP—ELISA法的建立

目的为改进端粒重复序列扩增法（ＴＲＡＰ）存在定量困难、应用同位素及每次检测标本数受限等缺点,研究建立及评价端粒酶ＴＲＡＰＥＬＩＳＡ法。方法端粒酶ＴＲＡＰＥＬＩＳＡ法是将ＴＲＡＰ与ＰＣＲＥＬＩＳＡ系统结合。与常规ＴＲＡＰ法相比较,应用端粒酶ＴＲＡＰＥＬＩＳＡ法检测端粒酶阳性的２９３细胞和阴性对照标本（加热或ＲＮａｓｅ处理和正常人内皮细胞）。结果２９３细胞端粒酶阳性,对１０,１０２,１０３及１０４个细胞检测均为阳性,所测到的吸光度值（Ａ,曾称光密度ＯＤ）依赖于被检２９３细胞数。ＲＮａｓｅ或加热处理标本和正常人内皮细胞均阴性。该方法可在当日观察结果,不需放射性同位素。结论端粒酶ＴＲＡＰＥＬＩＳＡ是一种非放射性同位素、快速及可定量的人端粒酶活性检测方法。     

Indomethacin and telomerase activity in tumor growth retardation

It is well-recognized that cycloogygenase inhibitors attenuate tumor growth in tumor models, although underlying mechanisms are unclear. In the present study we report that indomethacin retards MCG-101 tumor growth on mice by induction of apoptosis/necrosis and inhibits telomere elongation. The inhibition of telomerase activity by NSAIDs (indomethacin, mobic, sulindac sulfone, suramin) was, however, not a universal finding, since a mouse melanoma (K1735-M2) did not respond. By contrast, a human cell line of colon carcinoma origin (HT-29), responded by both retarded growth and telomerase activity despite a low intrinsic production of prostaglandins, mainly PGE2. Therefore, it is not likely that indomethacin inhibition of tumor growth and telomere elongation is directly related to Cox-1/Cox-2 activities in tumor cells. Also, NSAIDs at 25 microM (sulindac sulfone) decreased growth and telomerase activity in MCG-101 cells, without any effects on PGE2 production, while ibuprofen reduced PGE2 production but had no effect on growth or telomerase activity. Our results demonstrate that cyclooxygenase inhibitors can retard tumor growth both in murine tumors and in human tumor cells by inhibition of telomerase activity in addition to previously recognized mechanisms as induction of apoptosis, inhibition of cell proliferation, influence on the expression of growth factors around growing tumors and attenuation of neoangiogenesis.     

肺癌组织中端粒酶基因与端粒酶活性关系的研究

目的 探讨肺癌组织中端粒酶基因hTR、hTERT与端粒酶活性的表达是否与肺癌的发生发展有关，深入了解端粒酶基因对端粒酶活性的调控是在基因水平还是在转录水平。方法 采用TRAP－PCR和RT－PCR方法分别检测68例肺癌组织及相应癌旁肺组织中端粒酶活性、端粒酶基因hTR和hTERP的表达。结果 68例肺癌组织中端粒酶阳性率为79．4％（54／68）,hTR阳性率为98．5％（67／68）,hTERT阳性率为91．2％（62／68）。68例癌旁组织中无一例表达端粒酶阳性，但大多数癌旁组织均表达hTR(62/68,91.2%)，仅7例hTERT表达阳性（10．3％），与hTR相比，hTERT同端粒酶具有更高的一致性，其一致率为89．0％（121／136），而ThTR与端粒酶的一致率为43．4％（59／136）。结论 端粒酶的活性可能在肺癌发生发展中起重要的作用。hTR与hTERT可能是在转录后或翻译后水平对端粒酶进行调控的。     

乳腺癌组织中端粒酶活性的研究

目的细胞中端粒(telomere)的长度与细胞寿命的调控密切相关,端粒长度的维持需要端粒酶(telomerase)的激活,近年来的研究发现,端粒酶的激活与肿瘤的发生,发展关系密切.本文研究乳腺癌组织,癌旁组织,良性乳腺病变,正常乳腺组织中的端粒酶活性,探讨其作为乳腺癌肿瘤标志物的可能性.方法采用端粒重复序列扩增法(telomeraic repeat amplification protocol,TRAP)来检测36例乳腺癌及其相应癌旁组织,12例良性乳腺病变,6例正常乳腺组织中的端粒酶活性.结果 36例乳腺癌组织中,有33例端粒酶表达阳性,其阳性率为91.7%,而且与肿瘤的大小,淋巴结的状态,临床分期有相关性.36例癌旁组织中,有2例端粒酶表达阳性,阳性率为5.6%.12例良性乳腺病变中,仅有1例端粒酶表达阳性,阳性率为8.3%.6例正常乳腺组织端粒酶表达均为阴性.结论乳腺癌组织中普遍存在端粒酶活性表达,端粒酶有可能成为诊断乳腺癌的肿瘤标志物.     

大肠癌组织中端粒酶活性的研究

目的研究大肠癌组织及相应癌旁组织中的端粒酶活性,探讨其作为肠癌肿瘤标记物的可能性。方法采用ＴＲＡＰ方法研究了４０例大肠组织（包括３７例大肠癌,３例良性大肠疾病）和２０例相应癌旁上皮组织中的端粒酶活性表达。结果２０例癌旁上皮和３例良性疾病组织中,端粒酶活性表达均为阴性;而３７例大肠癌组织中,３５例显示端粒酶活性表达为阳性,阳性率为９４．６％。结论端粒酶是特异性较强的恶性肿瘤基因标志,有可能成为大肠癌早期诊断和治疗的理想标志物     

食管鳞癌组织端粒酶亚基的表达及端粒酶活性的关系

目的　探讨食管鳞癌组织中端粒酶活性、端粒酶逆转录酶 (h TERT)及端粒酶相关蛋白 - 1(TP- 1)的表达及其关系。方法　应用 TRAP-银染法对 45例食管鳞癌组织端粒酶活性的检测 ,同时应用原位杂交对癌组织切片进行 h TERT、TP- 1的 m RNA表达的检测。结果　癌组织端粒酶活性阳性率为 82 .2 %。癌组织中不同分化程度的端粒酶活性差异无显著性 (P>0 .0 5 )。有淋巴结转移者癌组织端粒酶活性明显高于无淋巴结转移者 ,差异有显著性(P<0 .0 5 )。癌组织中 h TERTm RNA表达的阳性率为 6 4.4%,TP- 1的阳性率为 6 2 .2 %。 h TERT的 m RNA表达与端粒酶活性密切相关 ,而 TP- 1的 m RNA表达与端粒酶活性无相关。结论　食管鳞癌组织中端粒酶活性及h TERT、TP- 1的 m RNA表达均较高。端粒酶活性与淋巴结转移有关。 h TERT与端粒酶活性有密切关系。     

DETECTION OF TELOMERASE ACTIVITY IN BREAST CARCINOMA

Ｔｅｌｏｍｅｒｅｓａｒｅｓｐｅｃｉａｌｉｚｅｄｓｔｒｕｃｔｕｒｅｓａｔｔｈｅｅｎｄｓｏｆｅｕｋａｒｙｏｔｉｃｃｈｒｏｍｏｓｏｍｅｓｗｈｉｃｈｆｕｎｃｔｉｏｎｉｎｍａｉｎｔａｉｎｉｎｇｃｈｒｏｍｏｓｏｍａｌｉｎｔｅｇｒｉｔｙ１Ｈｕｍａｎｔｅｌｏｍｅｒ...     

鼻咽癌组织端粒酶活性的研究

探讨端粒酶活性与鼻咽癌的关系。方法采用端粒重复序列扩增法测定６９例经病理确诊的鼻咽活检组织及两株鼻咽癌细胞株的端粒酶活性。     

DETECTION OF HUMAN TELOMERASE ACTIVITY BY TELOMERASE TRAP-ELISA ASSAY

ＤＥＴＥＣＴＩＯＮＯＦＨＵＭＡＮＴＥＬＯＭＥＲＡＳＥＡＣＴＩＶＩＴＹＢＹＴＥＬＯＭＥＲＡＳＥＴＲＡＰ—ＥＬＩＳＡＡＳＳＡＹＷｅｉＬｉｘｉｎ卫立辛ＷｕＭｅｎｇｃｈａｏ吴孟超ＹａｎＺｈｅｎｌｉｎ阎振林ＳｈｅｎＦｅｎｇ沈锋ＸｉｅＴｉａｎｐｅｉ谢天培Ｑｉａ...     

非小细胞肺癌端粒酶活性与端粒酶RNA、端粒酶催化亚单位的相关性及其意义

目的 探讨非小细胞肺癌（non-small cell lung cancer,NSCLC)端粒酶活性（telomerase activity,TA)、端粒酶RNA（telomerase RNA,hTR）端粒酶催化亚单位（telomerase catalytic subunit,hTRT/hEST2)编码基因的表达,彼此间相关性及其预后意义。方法 TRAP－PCR（telomerase repeat amplification protocol PCR)检测TA,RT－PCR检测hTR,原位杂交检测hTRT/hEST2mRNA表达。结果 非小细胞肺癌TA、hTR和hTRT/hEST2 mRNA阳性率分别为68．4％（26／38）、55．2％（32／58）和74．1％（43／58）,TA与hTRT/hEST2阳性相关（r=0.84,P=0.01),与hTR无相关性（r=0.16,P=0.23）。低分化以及有淋巴结或远处转移者TA及hTRT/hEST2阳性率增高;TA及hTRT/hEST2阳性组中位生存期（10．4个月,7．5个月）低于阴性组（13．5个月,14．7个月;P＝0．074,P＝0．01）,hTR阳性组中位生存期（12．9个月）略高于阴性组（11．5个月,P＝0．23）,仅hTRT/hEST2阳性与阴性组生存期差异有显著性;多因素Cox回归分析hTRT/hEST2对生存期有影响。结论 非小细胞肺癌TA、hEST2阳性与阴性组生存期差异有显著性;多因素Cox回归分析hTRT/hEST2对生存期有影响。结论 非小细胞肺癌TA、hTR和hTRT/hEST2表达高于癌旁正常组织;hTRT/hEST2够能反应TA活性,二者为NSCLC恶性表型增高的标志;hTRT/hEST2具有独立的预后意义。     

大肠癌端粒酶活性与c-myc基因表达的关系

目的　探讨端粒酶活性和c myc表达的相关性及其与大肠癌发生发展的关系。方法　运用端粒重复扩增 (TRAP)及免疫组织化学方法对 30例大肠癌、癌旁组织及正常粘膜和 2 0例大肠腺瘤性息肉组织中端粒酶活性和c myc蛋白进行检测。结果　大肠癌端粒酶活性和c myc蛋白表达率分别为 83 33% (2 5 / 30 )和 80 0 0 % (2 4 / 30 ) ,显著高于癌旁、正常大肠粘膜及大肠腺瘤性息肉组织 (P <0 0 5 ) ;端粒酶活性及c myc蛋白阳性表达率在伴淋巴结转移的大肠癌组织中明显高于无淋巴结转移者 (P <0 0 5 ) ;在 2 5例端粒酶活性阳性的大肠癌组织中有 2 3例 (92 0 0 % )表达c myc蛋白 ,端粒酶活性与c myc蛋白表达密切相关 (P <0 0 5 )。结论　端粒酶激活及c myc的异常表达与大肠癌的发生发展密切相关 ,联合检测有助于大肠癌的诊断和预后判断 ;在大肠癌的形成和发展过程中 ,c myc过度表达对端粒酶的激活可能起重要作用。     

Differential telomerase activity, expression of the telomerase catalytic sub-unit and telomerase-RNA in ovarian tumors.

Telomerase activity has been found in a variety of malignant tumors but only rarely in benign tumors or normal tissues. In this study, we investigated telomerase activation in 37 ovarian tumors, including benign, borderline and malignant neoplasms. Telomerase activity was detected using the telomeric repeat amplification protocol (TRAP) in 13/16 ovarian carcinomas, 9/10 borderline tumors and 3/11 cystadenomas/fibromas. mRNA expression of the putative human telomerase catalytic sub-unit gene (hTERT) was detected by RT-PCR in 14/15 ovarian carcinomas, 8/10 borderline tumors and 4/11 cystadenomas/fibromas. In situ hybridization was performed to evaluate telomerase-RNA (hTR) expression in the corresponding paraffin-embedded tumors. Variable expression levels of hTR were found over neoplastic tumor cells. The highest levels of hTR expression were found predominantly in ovarian carcinomas. Although the amount of telomerase activity varied, significantly high levels of telomerase activity were found predominantly in ovarian carcinomas. hTERT mRNA expression was closely associated with telomerase activity. These findings suggest that up-regulation of hTERT and hTR is important for telomerase activation during malignant-tumor progression. Telomerase activation might therefore be a valuable diagnostic parameter that could help to identify potentially progressive lesions. However, the diagnostic and therapeutic implications of telomerase activation need to be clarified in clinical trials. Int. J. Cancer (Pred. Oncol.) 84:426-431, 1999.     

肺癌组织中端粒酶活性检测及临床意义

目的　研究肺癌组织中端粒酶活性检测的临床意义 ,探讨其作为肺癌诊断和预后评价指标的可行性。方法　采用端粒重复序列扩增 (telomericrepeatamplificationprotocal ,TRAP) PCR ELISA方法检测 48例肺癌手术切除组织及相应的癌旁组织中端粒酶的活性 ,并以 42例肺部良性病变组织作为对照。结果　 48例肺癌组织中端粒酶阳性率为 87.5 % ( 4 2 /4 8) ,癌旁组织为 8.3 % ( 4 /4 8) ,42例肺部良性病变组织中未检出端粒酶阳性。肺癌组织端粒酶水平明显高于癌旁组织 ( χ2 =13 .0 2 9,P <0 .0 1)及良性病变组织 ( χ2 =14 .0 16,P <0 .0 1)。端粒酶活性随细胞分化程度、临床分期有升高趋势 ,但无统计学差异 ,不同组织学类型间端粒酶活性无显著性差异 ,伴淋巴结转移肺癌组织中端粒酶阳性率明显高于不伴淋巴结转移组 ( 93 .5 %比 76.4% ,χ2 =63 .5 11,P <0 .0 1)。结论　端粒酶的激活与肺癌的发生发展密切相关。端粒酶可作为肺癌辅助诊断、预后判断的标志物之一 ,并为肺癌的基因治疗提供理论依据     

Clinical significance of telomerase activity in multiple myeloma

BACKGROUND: The clinical course of patients with multiple myeloma varies, and therefore it is important to evaluate the disease state. We studied the telomerase activity of myeloma cells as a possible prognostic factor in such patients. METHODS: Twenty five samples from patients with multiple myeloma were studied. We purified myeloma cells in bone marrow samples according to the expression of surface antigens, CD38 and CD45. CD38+/CD45- or dim cells had morphologic characteristics of myeloma cells, with a purity exceeding 95%. The telomerase activity of myeloma cells was determined by a polymerase chain reaction-based telomeric repeat amplification protocol assay. Ki-67 positivity of the purified cells was determined by flow cytometry using anti-Ki-67 antibody. The relationship between telomerase activity and prognostic factors was also examined. RESULTS: A significantly high degree of telomerase activity was detected in subjects with a serum beta2-microglobulin level > or = 6 mg/dL or at Stage III (P = 0.002). The serum C-reactive protein, lactate dehydrogenase, and creatinine levels did not correlate with the telomerase activity, but this activity did significantly correlate with Ki-67 positivity and the percentage of plasma cells in the bone marrow (r = 0.561, P = 0.004, and r = 0.397, P = 0.049, respectively). The patients with high levels of telomerase activity were thus found to have a significantly short survival time after sampling (P = 0.035). CONCLUSIONS: The measurement of the telomerase activity in myeloma cells was found to be a reliable marker for the proliferating capacity and tumor mass in myeloma patients. The telomerase activity of myeloma cells may therefore be useful as a prognostic factor.